title: A fraction of stem bark extract of Entada africana suppresses lipopolysaccharide-induced inflammation in RAW 264.7 cells.
abstract: BACKGROUND
  Entada africana is a plant used in African traditional medicine for the treatment of stomachache, fever, liver related diseases, wound healing, cataract and dysentery.
  
  
  OBJECTIVE
  This study aimed at evaluating the anti-inflammatory activity of fractions of the stem bark extract of the plant using lipopolysaccharide (LPS)-induced inflammation in RAW 264.7 macrophages model.
  
  
  METHODS
  The crude extract was prepared using the mixture CH2Cl2/MeOH (1:1, v/v) and fractionated by flash chromatography using solvents of increasing polarity to obtain five different fractions. The effects of the fractions on the cells viability were studied by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and their inhibitory activity against LPS-induced nitric oxide (NO) production screened by Griess test. The most active fraction was further investigated for its effects on reactive oxygen species (ROS) production using flux cytometry, the expression of inducible nitric oxide synthase (iNOS), pro-and anti-inflammatory cytokines (IL1β, TNFα, IL6, IL10 and IL13) by RT-PCR, and the activity of the enzyme p38 MAPK kinase by enzyme-linked immunosorbent assay (ELISA).
  
  
  RESULTS
  The fractions presented no significant effect on the viability of macrophages at 100 μg/ml after 24h incubation. The CH2Cl2/MeOH 5% (Ea5) fraction was found to be the most potent in inhibiting NO production with a half inhibition concentration (IC50)=18.36 μg/ml, and showed the highest inhibition percentage (89.068%) in comparison with Baicalin (63.34%), an external standard at 50 μg/ml. Ea5, as well as Baicalin significantly (P<0.05) inhibited the expression of TNFα, IL6 and IL1β mRNA, attenuated mRNA expression of inducible NO synthase in a concentration-dependent manner, stimulated the expression of anti-inflammatory cytokines (IL10 and IL13), and showed a 30% inhibition of the activity of p38 MAPK kinase.
  
  
  CONCLUSIONS
  The results of the present study indicate that the fraction Ea5 of Entada africana possesses most potent in vitro anti-inflammatory activity and may contain compounds useful as a therapeutic agent in the treatment of inflammatory related diseases cause by over-activation of macrophages.
authors.affiliation: Division of Immunopathology of Nervous System, Department of Neuropathology, Institute of Pathology, University of Tübingen, Germany. briceayissi03@yahoo.fr
authors.forename: Brice
authors.initials: B
authors.lastname: Ayissi Owona
authors.forename: Nico Frederic
authors.initials: NF
authors.lastname: Njayou
authors.forename: Stefan
authors.initials: S
authors.lastname: Laufer
authors.forename: Paul Fewou
authors.initials: PF
authors.lastname: Moundipa
authors.forename: Hermann J
authors.initials: HJ
authors.lastname: Schluesener
mesh_terms: D000818:Animals; D000893:Anti-Inflammatory Agents / Q000302:isolation & purification / Q000494:pharmacology*; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D016207:Cytokines / Q000276:immunology; D004305:Dose-Response Relationship, Drug; D020834:Ethnopharmacology; D007887:Fabaceae / Q000737:chemistry*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects* / Q000276:immunology / Q000473:pathology; D016488:Medicine, African Traditional; D051379:Mice; D009569:Nitric Oxide / Q000096:biosynthesis; D024301:Plant Bark / Q000737:chemistry; D010936:Plant Extracts / Q000302:isolation & purification / Q000494:pharmacology*; D018547:Plant Stems / Q000737:chemistry; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D000893:Anti-Inflammatory Agents; D016207:Cytokines; D008070:Lipopolysaccharides; D010936:Plant Extracts; D017382:Reactive Oxygen Species; D009569:Nitric Oxide
keywords: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; Baicalin; Cytokines; Ea5; Entada africana; Entada africana fraction CH(2)CL(2)/MeOH 5%; MAPK; MTT; Macrophages; NO; ROS; iNOS; inducible nitric oxide synthase; mitogen activated protein kinase; nitric oxide; p38 MAPK kinase; reactive oxygen species